P. aeruginosa biofilms cause many chronic infections including airway infections that afflict bronchiectasis patients, endotracheal tube colonization that leads to ventilator associated pneumonia, medical device infections, and others. In biofilms, bacteria live in matrix-encased group structures, and this growth mode produces physiologic changes that cause marked antibiotic resistance. In spite of the importance of biofilms in disease, no medical treatments have yet been developed to disrupt them. Interestingly, biofilm bacteria themselves possess active mechanisms to separate from biofilms; this process is known as detachment. Because detached bacteria regain sensitivity to killing, this process may be useful therapeutically. The goal of this work is to investigate a potential mechanism of P. aeruginosa biofilm detachment, and to study the pathological consequences of detachment using an airway biofilm infection model. We have discovered a P. aeruginosa variant strain with a hyper-detachment phenotype. This strain overproduces the bio-surfactant, rhamnolipid, which is required for its accelerated biofilm detachment. The following aims are proposed: 1. What is the mechanism of rhamnolipid-mediated biofilm detachment? We will examine the types of rhamnolipids produced by the wild type and variant strain, and use two independent methods to determine if rhamnolipids can, by themselves, cause biofilm detachment. Microarray analysis will be used to find other genes that may be needed for detachment. 2. What are the consequences of biofilm detachment in vivo? Using our airway biofilm infection model and isogenic hyper and hypo-detaching strains, we will investigate the potential risks of inducing detachment and determine if detachment improves clearance of the organisms by the host. 3. Will biofilm detachment in vivo increase the efficacy of antibiotic treatment? Using non-detaching strains, infections in the airway biofilm model are extremely resistant to treatment. In vitro, biofilm detachment restores antibiotic sensitivity. We will investigate whether detachment increases the efficacy of antibiotic treatment in vivo. These studies will model a treatment strategy that combines traditional antibiotics with induced biofilm detachment. [unreadable] [unreadable]